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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The bovine 17 alpha-hydroxylase cytochrome P450 gene (CYP17) contains at least two cAMP-responsive sequences (CRS) within its 5'-flanking region. In this study it is demonstrated that one of the sequences, CRS1, is also a target for protein kinase C (PKC)-mediated regulation. Forskolin-induced, CRS1-dependent transcription of a heterologous minimal promoter/structural gene which had been transfected into the mouse adrenocortical tumor cell line Y1 was suppressed by activation of PKC by phorbol esters such as 12-O-tetradecanoyl phorbol-14-acetate and phorbol 12,13-didecanoate-beta (PDD beta). Use of the active and inactive forms of PDD (PDD alpha and PDD beta) as well as down-regulation of PKC by prolonged treatment of the cells with 12-O-tetradecanoyl phorbol-14-acetate demonstrated that the effect of phorbol esters on transcription conferred by CRS1 was mediated through the PKC pathway and not a consequence of general toxicity to the cells. Analysis of the different steps in the signal transduction pathway between the adenylate cyclase and the CRS1 element suggests that phrobol esters do not exert their effect by altering the forskolin-induced cAMP production, activation of PKA, or the binding of nuclear proteins to CRS1. These results establish the CRS1 element as a target not only for PKA, but also for the PKC-mediated signal transduction pathway. They further suggest that PKC interferes with the transcriptional activation competence of factors bound to CRS1 and the minimal promoter.
Mol Endocrinol 1992 Aug
PMID:A novel 3',5'-cyclic adenosine monophosphate-responsive sequence in the bovine CYP17 gene is a target of negative regulation by protein kinase C. 132 75

There are at least three isozymes (C alpha, C beta, and C gamma) of the mammalian catalytic (C) subunit of cAMP-dependent protein kinase (PKA) (Beebe, S., Oyen, O., Sandberg, M., Froysa, A., Hansson, V., and Jahnsen, T. (1990) Mol. Endocrinol. 4, 465-475). To compare the C gamma and C alpha isozymes, the respective cDNAs were expressed in permanently transformed Kin-8 PKA-deficient Y1 adrenal cells using the mouse metallothionein promoter. The recombinant C subunits were characterized as immunoreactive, zinc-inducible, cAMP-dependent kinase activities. In contrast to C alpha, histone was a better substrate than Leu-Arg-Arg-Ala-Ser-Leu-Gly (Kemptide) for C gamma. Furthermore, C gamma histone kinase activity was not inhibited by the protein kinase inhibitor peptide (5-24 amide), which has been widely used as a PKA-specific inhibitor. The major C gamma peak (type I) eluted from DEAE-Sepharose at a higher NaCl concentration (120 mM) than the C alpha type I eluted (70 mM). C gamma and C alpha type II eluted between 220 and 240 mM NaCl. C gamma required higher concentrations of cAMP than C alpha did for dissociation from the mutant type I holoenzyme. These differences provided a basis for the separation of the mutant RI-associated isozymes on DEAE-Sepharose. Both C alpha (41-42 kDa) and C gamma (39-40 kDa) were identified by a C subunit antibody after sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblot analysis. Zinc induced the PKA-mediated rounding phenotype in C gamma and C alpha clones, thereby restoring the cells to the parent Y1 adrenal cell phenotype. Collectively, these data indicate that C gamma is an active PKA C subunit but suggest that C gamma and C alpha have different protein and peptide recognition determinants.
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PMID:The C gamma subunit is a unique isozyme of the cAMP-dependent protein kinase. 133 96

Protein kinase A (PKA) has been shown to modulate the pattern of gene expression via transcription factors such as cAMP response element binding protein. However, in F9 embryonal carcinoma cells which lack endogenous functional cAMP response element binding protein, we have found that PKA is still able to control gene transcription through the glucocorticoid receptor (GR) by up-regulating its hormone-dependent trans-activation. Dose-response analysis indicates that PKA does not alter the ligand binding affinity of GR. PKA seems to act through the DNA binding domain of GR, since GR mutants which lack either the amino-terminal or the ligand binding domain are still able to be up-regulated by PKA. In support of this proposal, we demonstrate that PKA can enhance the DNA binding activity of GR. Our results suggest a novel mechanism by which PKA modulates the steroid sensitivity of a target cell by enhancing the DNA binding activity of GR for its cognate hormone response elements.
Mol Endocrinol 1992 Sep
PMID:Modulation of glucocorticoid receptor function by protein kinase A. 143 89

The morphological and functional characteristics and the activities of cyclic AMP- (PKA I and PKA II) and calcium and phospholipid-dependent (PKC) protein kinases were studied in 2-day-old suspension cultures of porcine thyroid cells and were compared with those in freshly dissociated cells and intact glands. Thyroid cell morphology changed during the 2-day culture in the absence of specific regulators. This is characterized by a loss of cellular polarity, exo- and endocytotic vesicles and membranes of the rough endoplasmic reticulum, and an increase in the number of lysosomes, pseudomyelinic structures, lipidic inclusions and free ribosomes. Functional changes are characterized by a progressive decrease in protein iodination and its sensitivity to TSH stimulation. The total PKA activity in the cytosols of these cultures was slightly greater than that of freshly prepared tissue, due to the selective and significant accumulation of PKA I in cultured cells. In the particulate fraction the PKA activity was unchanged. PKC is the major kinase activity in porcine thyroids, and remains so in cultured cells. The slight drop in its activity in cytosols was offset by a significant increase in the particulate fraction, suggesting an intracellular redistribution of this kinase in cultured cells. The PKC activity is also partly activated in both the cytosol and particulate fraction, which results in an increased basal activity. The changes in PKA and PKC activities greatly modified the PKC/PKA ratios in the cytosols and the particulate fractions of cultured cells. These modifications could be partly responsible for the changes in sensitivity of cultured cells to the agents which control their activity.
Mol Cell Endocrinol 1990 Jul 09
PMID:Changes in cAMP-dependent and Ca2(+)-phospholipid-dependent protein kinase activities in suspension cultures of porcine thyroid cells. 217 Feb 12

Protein kinase A (cAMP-dependent) and C (calcium, phospholipid-dependent) activities were measured and in vitro phosphorylation of endogenous proteins by these kinases were observed by SDS-PAGE in 100,000 x g supernatant (soluble) fractions of ovine small (12-22 microns) and large (greater than 22 microns) luteal cells. No differences in stimulation (P less than 0.05) of A kinase activity between small and large cells were detected. Protein kinase C activity was stimulated (P less than 0.05) 2.9-fold in small cells but not significantly enhanced above basal (P greater than 0.05) in large cells. By direct comparison, greater stimulation (P less than 0.05) over basal of A versus C kinase (6.1- versus 2.9-fold) was measured in small cells. These stimulations were greater than those observed in large cells (A kinase, 4.8-fold; C kinase, 1.8-fold). Maximal specific activities of both kinases (per mg protein) were greater (P less than 0.05) in small than in large cells. Endogenous proteins that could serve as substrates for phosphorylation by A and C kinases differed between small and large cells. Phosphorylation of six proteins by A kinase was consistently greater in small than in large cells. One endogenous protein (37 kDa) appeared to serve as a preferred substrate for phosphorylation by A kinase in small cells and C kinase in large cells. One protein (81 kDa) was predominantly phosphorylated in large rather than small cells by a calcium-dependent, C kinase-independent mechanism. These results support the accepted role of cAMP via A kinase and a possible role for C kinase in regulating steroidogenesis in ovine small luteal cells. The inability of large cells to respond to cAMP with enhanced secretion of progesterone may be due to an unavailability of phosphoprotein substrates for A kinase. Furthermore, protein kinase C activity and available protein substrates display quantitative and qualitative differences between small and large cells. Differences in regulation of steroidogenesis between the cell types may be due to these observed differences.
Mol Cell Endocrinol 1989 Apr
PMID:Protein kinase A and C activities and endogenous substrates in ovine small and large luteal cells. 254 89

Protein kinase A (PKA) dependent phosphorylation of C-protein and cardiac troponin I (cTnI) is known to be associated with a reduced sensitivity to Ca2+. We have investigated the relative importance of each of these sites of phosphorylation in this effect by use of extraction/reconstitution experiments and mutagenesis of recombinant cTnI. Conditions developed for extraction of troponin (Tn) complex also resulted in extraction of C-protein. A truncated cTnI (cTnI/NH2) lacking the 32 amino acids in the unique amino terminal extension of cTnI was engineered and expressed. In contrast to native cTnI, cTnI/NH2, which lacks Ser23 and Ser24, was not phosphorylated by PKA either in pure form or after incorporation into the myofilament lattice. The relation between pCa (-log molar free Ca2+ concentration) and MgATPase activity of non-phosphorylated native myofibrils or non-phosphorylated myofibrils reconstituted with cTnI, but lacking C-protein, was the same and could not be distinguished from that of control or PKA-treated myofibrils into which we exchanged cTnI with cTnI/NH2. However, PKA-dependent phosphorylation of either native myofibrils or reconstituted myofibrils containing cTnI but lacking C-protein resulted in an identical and significant rightward shift of pCa50 (half-maximally activating pCa) in the pCa-activity relationship. Our results strongly indicate that phosphorylation of cTnI at Ser residues in the unique amino terminal extension of the molecule is both necessary and sufficient for the decrease in myofilament Ca(2+)-sensitivity associated with PKA-dependent phosphorylation.
J Mol Cell Cardiol 1995 Jul
PMID:The unique amino-terminal peptide of cardiac troponin I regulates myofibrillar activity only when it is phosphorylated. 747 84

Neuropeptide Y (NPY) is a 36 amino acid peptide present in the central and peripheral nervous systems. Treatment with Nerve Growth Factor (NGF) induces an increase in NPY mRNA in PC12 cells, a rat pheochromocytoma cell line extensively used as a model of neuronal differentiation. Stimulators of both cAMP and calcium-phospholipid dependent protein kinases (PKA and PKC respectively) increase NPY mRNA levels in a similar way to NGF. Nevertheless, H-89, a specific inhibitor of PKA failed to block NGF stimulated NPY mRNA accumulation. Furthermore, direct measurement of PKA activity in cell extracts showed no increase following NGF, in contrast to forskolin. H7, an inhibitor of both PKC and PKA systems completely abolished the NGF induced increase in NPY mRNA, suggesting that PKC is necessary for NGF induction of the NPY gene. NGF also increased PKC activity in cell extracts in a similar way to phorbol myristate acetate (PMA). Use of a reporter function, chloramphenicol acetyl transferase, controlled by 700 base pairs of the 5' flanking region of the NPY gene demonstrated that NGF and phorbol ester stimulated transcription of the NPY gene. This stimulation could be blocked by pre-incubating PC12 cells with calphostin C, a specific inhibitor of PKC. Our results indicate that NGF induces NPY gene expression via activation of PKC system. Although an increase in adenylate cyclase activity affects the expression of the NPY gene, activation of PKA appears not to be involved in mediating the NGF effects.
Brain Res Mol Brain Res 1994 Jun
PMID:Role of protein kinase C in mediating NGF effect on neuropeptide Y expression in PC12 cells. 752 99

Differential expression of PAI-1 in connective tissues has been associated etiologically with some forms of arthritis. Our objective was to delineate the mechanisms by which PGE2 and IL-1 beta, inflammatory mediators commonly found at sites of inflammation, regulate the expression and synthesis of PAI-1 in human synoviocytes. PGE2 (and PGE1) inhibited PAI-1 mRNA expression and secretion in a dose-dependent manner with an IC50 (for antigen secretion) of 4.6 x 10(-10) M and 8.7 x 10(-10) M, respectively. Cyclic AMP agonists forskolin, Sp-cAMP, and IBMX mimic the effects of the PGEs. rhIL-1 beta stimulated the secretion of PAI-1 in a dose-dependent fashion under basal culture conditions; the effect was reversed by actinomycin D and the protein kinase inhibitors H7 and staurosporine but not KT-5720. PMA, an activator of protein kinase C, transiently increased (maximum 3 h) the expression of PAI-1 mRNA by approximately 10-fold, especially the 3.2 kb species. However, there was no significant increase in PAI-1 antigen secreted into the culture medium after PMA (100-300 nM) treatment. The half-life (t1/2) of PAI-1 mRNA, both the 3.2 and 2.2 transcripts was about 9.6 h (mean n = 3) and PGE2 has no affect on the stability of both messages. PGE2 reduced the rate of PAI-1 gene transcription as judged by run-off assays. The NSAID naproxen (30 micrograms/ml) induced the expression of PAI-1 mRNA over basal levels and super-induced the inhibitor's expression above rhIL-1 beta stimulated levels. Our results suggest that PGE2 suppresses PAI-1 expression and synthesis by activation of the cAMP/PKA system and inhibition of the rate of gene transcription. Data concerning the activation of PKC suggest that the expression, synthesis and release of the PAI-1 may be differentially regulated in normal human synoviocytes.
Mol Cell Endocrinol 1994 Jul
PMID:Transcriptional regulation of plasminogen activator inhibitor-1 expression in human synovial fibroblasts by prostaglandin E2: mediation by protein kinase A and role of interleukin-1. 752 83

In the brain, the 5 alpha-reductase converting testosterone (T) is present both in neurons and in glial cells, even if it prevails in neurons; the 3 alpha-hydroxysteroid-dehydrogenase (3 alpha-HSD), the enzyme converting dihydrotestosterone (DHT) into 3 alpha-diol, is particularly concentrated in type 1 astrocytes. In glial cells, since the 5 alpha-reductase is activated by a cAMP analogue, PKA seems to be involved in the control of this enzyme, postulating that nervous inputs utilizing cAMP as the second messenger might modify the activity of this enzyme in glial cells. Moreover, the results indicate that, in type 1 astrocytes, both the 5 alpha-reductase and the 3 alpha-HSD are stimulated by the co-culture with neurons and by the addition of neuron-conditioned medium, suggesting that secretory products released by neurons might intervene in the control of glial cell function.
J Steroid Biochem Mol Biol 1995 Jun
PMID:Metabolism of steroids in pure cultures of neurons and glial cells: role of intracellular signalling. 762 76

The catalytic subunit of turtle (Trachemys scripta elegans) liver cyclic AMP-dependent protein kinase (PKAc) was purified to homogeneity with a final specific activity of 65,783 pmol phosphate transferred.min-1.mg protein-1. Subunit molecular weight was 42-43 kDa as determined by SDS-PAGE and Sephacryl S-300 chromatography. The isolectric point was pH 6.41 +/- 0.02. Turtle liver PKAc showed highest activity with kemptide as its substrate; activity with other artificial substrates, histone IIA and protamine, was only 21 and 11%, respectively, of the activity with kemptide. Km values were 83 +/- 6.5 microM for Mg.ATP and 11.7 +/- 0.5 microM for kemptide and enzyme activity was strongly reduced by inhibitors of mammalian PKA (H-89, PKA-1) but not by inhibitors of other protein kinases. The enzyme was also inhibited by salts, especially fluoride salts (I50 about 30 mM), and showed a sharp break in the Arrhenius plot (0-45 degrees C) with activation energy increasing by 4-fold from 27.9 +/- 1.85 to 115 +/- 2.5 kJ/mol for temperatures above versus below 15 degrees C. Temperature effects may be important in suppressing PKA function, and therefore PKA-mediated responses, in vivo to enhance anoxic survival time during winter hibernation under water. Analysis of the effects of in vivo anoxia exposure at 7 degrees C on PKA in turtle organs showed a rapid 2.3-fold increase in the amount of active enzyme in liver within 1 h of anoxic submergence accompanied by a 60% increase cAMP levels; with longer anoxia (5 or 20 h) the percentage of active PKA was suppressed to 2.1-3.7% of the total.(ABSTRACT TRUNCATED AT 250 WORDS)
Mol Cell Biochem 1995 Apr 12
PMID:cAMP-dependent protein kinase and anoxia survival in turtles: purification and properties of liver PKA. 765 81


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